DESCRIPTION: (Applicant's Description) The relationship between the chemistry of ATP hydrolysis and the mechanical events that result in rotation of the gamma subunit of the F1-ATPase will be examined. Catalytic function of F1 derives from the asymmetry of the catalytic sites that, in turn, depends on the gamma subunit and the Mg2+ cofactor. The driving force for gamma rotation is believed to result from the initial binding energy of the Mg2+-ATP complex and from the release of phosphate which is a Mg2+ ligand. The ability of the gamma subunit of F1 to rotate will be measured using a fluorescent microsphere attached to the gamma subunit as recorded using a CCD camera. The torque generated during gamma rotation as well as the dwell time between rotations will be as assessed with F1 that contains site-directed mutants or other treatments at locations that may affect the coupling between hydrolysis and gamma rotation. Three loci are targeted for investigation that include: (a) Switch 3, the gamma subunit C-terminus and the beta subunit greasy bearing which is close to the site of Mg2+ binding and phosphate release; (b) Switch 2, the interface between the gamma subunit and the beta subunit DELSEED sequence; and (c) Switch 1, where hydrogen bonds form between the gamma subunit and the betaE subunit catch loop. Experiments will examine the possibility that Switch 1 is part of an escapement mechanism that only allows gamma rotation when the catalytic sites are filled with metal-nucleotide complex. Experiments are also designed to identify changes in metal ligands at the catalytic sites that are specifically associated with conformational changes linked to gamma rotation.